Chemical preconditioning prevents paradoxical increase in glutamate release during ischemia by activating ATP-dependent potassium channels in gerbil hippocampus

Ischemic tolerance induced by pretreatment with a low dose of 3-nitropropionic acid (3-NPA), called chemical preconditioning, prolongs the delay to hypoxic depolarization and improves the recovery of synaptic transmission (Exp. Neurol. 166 (2000), 385-391). We studied the effect of chemical preconditioning on the presynaptic site by analyzing spontaneous excitatory postsynaptic currents (sEPSCs) and miniature EPSCs (mEPSCs) with a whole cell patch-clamp technique in gerbil hippocampal slices. The frequency of sEPSCs decreased first and then dramatically increased during ischemia (10 min in duration, low pO(2), and deprivation of glucose) up to 200-300%. This increase was apparently a paradox, since synaptic transmission evoked by electrical stimulation diminished when the sEPSC frequency started to increase. The frequency of mEPSCs also increased in the same time course. Increases in sEPSC and mEPSC frequencies were prevented by chemical preconditioning with 3-NPA (4 mg/kg) administered intraperitoneally 3 h before the preparation of brain slices. These effects of chemical preconditioning were abolished by glibenclamide (5 microM), a blocker of ATP-dependent potassium (K(ATP)) channels, applied in vitro before the ischemic insult. The application of diazoxide (500 microM), an opener of K(ATP) channels, produced the same preventive effects on sEPSC and mEPSC frequencies. These results suggested that chemical preconditioning acted on presynaptic terminals to prevent the paradoxical increase in glutamate release during ischemia through the activation of K(ATP) channels.     

Long-term potentiation of primary afferent neurotransmission at trigeminal synapses of juvenile rats

Primary afferent monosynaptic and polysynaptic excitatory postsynaptic currents (EPSCs) were recorded from brainstem trigeminal neurons by stimulation of the mandibular nerve attached to the brainstem preparation of juvenile rats. A high-frequency conditioning stimulus induced long-term potentiation (LTP) of high-threshold EPSCs in the majority of trigeminal caudal neurons in substantia gelatinosa, where both A- and C-fibres terminate. However, the same conditioning stimulus did not potentiate low-threshold EPSCs in caudal neurons or EPSCs recorded from neurons in the middle part of trigeminal interpolar nucleus, where C-fibres rarely terminate. LTP in caudal neurons could be induced after blocking N-methyl-D-aspartate (NMDA) receptors with D(-)-2-amino-5-phosphonopentanoic acid (D-AP5, 50 microM), after postsynaptic loading of the Ca2+ chelator BAPTA (10 mM), or even after completely blocking excitatory transmission with kynurenic acid during conditioning. However, LTP was blocked by the metabotropic glutamate receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine (1 mM). We suggest that LTP of the trigeminal primary afferent EPSCs is induced preferentially in the C-fibre inputs and that the induction mechanism involves metabotropic glutamate receptors, possibly at the presynaptic terminals.     

Melatonin receptor activation increases glutamatergic synaptic transmission in the rat medial lateral habenula

Melatonin (MLT) is secreted from the pineal gland and mediates its physiological effects through activation of two G protein‐coupled receptors, MT₁ and MT₂. These receptors are expressed in several brain areas, including the habenular complex, a pair of nuclei that relay information from forebrain to midbrain and modulate a plethora of behaviors, including sleep, mood, and pain. However, so far, the precise mechanisms by which MLT control the function of habenula neurons remain unknown. Using whole cell recordings from male rat brain slices, we examined the effects of MLT on the excitability of medial lateral habenula (MLHb) neurons. We found that MLT had no significant effects on the intrinsic excitability of MLHb neurons, but profoundly increased the amplitude of glutamate–mediated evoked excitatory post‐synaptic currents (EPSC). The increase in strength of glutamate synapses onto MLHb neurons was mediated by an increase in glutamate release. The MLT‐induced increase in glutamatergic synaptic transmission was blocked by the competitive MT₁/MT₂ receptor antagonist luzindole (LUZ). These results unravel a potential cellular mechanism by which MLT receptor activation enhances the excitability of MLHb neurons. The MLT‐mediated control of glutamatergic inputs to the MLHb may play a key role in the modulation of various behaviors controlled by the habenular complex. Synapse, 2016. © 2016 Wiley Periodicals, Inc. Synapse 70:181–186, 2016. © 2016 Wiley Periodicals, Inc.     

Syntaxin 1A occludes GABA(B) receptor-induced inhibition of exocytosis downstream of Ca(2+) entry in mouse hippocampal neurons

The mechanisms underlying gamma-amino butyric acid (GABA(B)) receptor-mediated inhibition of exocytosis have been characterized in a variety of synapses. Using patch-clamp recording methods, we attempted to clarify the intracellular mechanisms underlying presynaptic inhibition in autaptic synapses of isolated mouse hippocampal neurons. Baclofen, a selective GABA(B) receptor agonist, decreased the frequency of glutamatergic miniature excitatory postsynaptic currents (mEPSCs) without changing their amplitude in Ca(2+)-free extracellular solution, suggesting that baclofen inhibits exocytosis downstream of Ca(2+) entry. Syntaxin 1A is known to modulate exocytosis and suppress neuronal sprouting. Antisense oligonucleotide-mediated knockdown of syntaxin 1A increased the frequency of mEPSCs under Ca(2+)-free condition. Estimation of the number of functional release sites by staining with FM1-43 indicated that the increased frequency of mEPSCs was induced by facilitation of exocytosis at each site, rather than by an increased number of release sites due to neuronal sprouting. Baclofen reduced mEPSC frequency in syntaxin 1A-knockdown neurons to the same level as that in nonsense oligonucleotide transfected neurons under Ca(2+)-free condition. These results suggest that the GABA(B) receptor- and syntaxin 1A-induced inhibitions of exocytosis occlude one another and that the GABA(B) receptor shares a common intracellular pathway with syntaxin 1A in inhibiting transmitter release downstream of Ca(2+) entry.     

Synaptic transmission changes in fear memory circuits underlie key features of an animal model of schizophrenia

Non-competitive antagonists of the N-methyl-d-aspartate receptor (NMDA) such as phencyclidine (PCP) elicit schizophrenia-like symptoms in healthy individuals. Similarly, PCP dosing in rats produces typical behavioral phenotypes that mimic human schizophrenia symptoms. Although schizophrenic behavioral phenotypes of the PCP model have been extensively studied, the underlying alterations of intrinsic neuronal properties and synaptic transmission in relevant limbic brain microcircuits remain elusive. Acute brain slice electrophysiology and immunostaining of inhibitory neurons were used to identify neuronal circuit alterations of the amygdala and hippocampus associated with changes in extinction of fear learning in rats following PCP treatment. Subchronic PCP application led to impaired long-term potentiation (LTP) and marked increases in the ratio of NMDA to 2-amino-3(5-methyl-3-oxo-1,2-oxazol-4-yl)propionic acid (AMPA) receptor-mediated currents at lateral amygdala (LA) principal neurons without alterations in parvalbumin (PV) as well as non-PV, glutamic acid decarboxylase 67 (GAD 67) immunopositive neurons. In addition, LTP was impaired at the Schaffer collateral to CA1 hippocampal pathway coincident with a reduction in colocalized PV and GAD67 immunopositive neurons in the CA3 hippocampal area. These effects occurred without changes in spontaneous events or intrinsic membrane properties of principal cells in the LA. The impairment of LTP at both amygdalar and hippocampal microcircuits, which play a key role in processing relevant survival information such as fear and extinction memory concurred with a disruption of extinction learning of fear conditioned responses. Our results show that subchronic PCP administration in rats impairs synaptic functioning in the amygdala and hippocampus as well as processing of fear-related memories.     

Localization and function of pre- and postsynaptic kainate receptors in the rat globus pallidus

Kainate receptors (KARs) are widely expressed the basal ganglia. In this study, we used electron microscopic immunocytochemistry and whole-cell recording techniques to examine the localization and function of KARs in the rat globus pallidus (GP). Dendrites were the most common immunoreactive elements, while terminals forming symmetric or asymmetric synapses and unmyelinated axons comprised most of the presynaptic labeling. To determine whether synaptically released glutamate activates KARs, we recorded excitatory postsynaptic currents (EPSCs) in the GP following single-pulse stimulation of the internal capsule. 4-(8-Methyl-9H-1,3-dioxolo[4,5 h]{2,3}benzodiazepine-5-yl)-benzenamine hydrochloride (GYKI 52466, 100 microm), an alpha-amino-3-hydroxyl-5-methyl-4-isoxazole propionic acid (AMPA) receptor antagonist, reduced but did not completely block evoked EPSCs. The remaining EPSC component was mediated through activation of KARs because it was abolished by 6-cyano-7-nitroquinoxaline-2, 3-dione (CNQX), an AMPA/KAR antagonist. The rise time (10-90%) and decay time constant (tau) for those EPSCs were longer than those of AMPA-mediated EPSCs recorded before GYKI 52466 application. KAR activation inhibited EPSCs. This inhibition was associated with a significant increase in paired-pulse facilitation ratio, suggesting a presynaptic action of KAR. KAR inhibition of EPSCs was blocked by the G-protein inhibitor, N-ethylmaleimide (NEM), or the protein kinase C (PKC) inhibitor calphostin C. Our results demonstrate that KAR activation has dual effects on glutamatergic transmission in the rat GP: (1) it mediates small-amplitude EPSCs; and (2) it reduces glutamatergic synaptic transmission through a presynaptic G-protein coupled, PKC-dependent, metabotropic mechanism. These findings provide evidence for the multifarious functions of KARs in regulating synaptic transmission, and open up the possibility for the development of pharmacotherapies to reduce the hyperactive subthalamofugal projection in Parkinson's disease.     

Enaminones and norepinephrine employ convergent mechanisms to depress excitatory synaptic transmission in the rat nucleus accumbens in vitro

We recently reported that anticonvulsant anilino enaminones depress excitatory postsynaptic currents (EPSCs) in the nucleus accumbens (NAc) indirectly via gamma-aminobutyric acid (GABA) acting on GABA(B) receptors [S.B. Kombian et al. (2005)Br. J. Pharmacol., 145, 945-953]. Norepinephrine (NE) and dopamine (DA), both known to be involved in seizure disorders, also depress EPSCs in this nucleus. The current study explored a possible interaction between enaminones and adrenergic and/or dopaminergic mechanisms that may contribute to their synaptic depression and anticonvulsant effect. Using whole-cell recording in rat forebrain slices containing the NAc, we show that NE-induced, but not DA-induced, EPSC depression occludes E139-induced EPSC depressant effect. UK14,304, a selective alpha(2) receptor agonist, mimicked the synaptic effect of NE and also occluded E139 effects. Phentolamine, a non-selective alpha-adrenergic antagonist that blocked NE-induced EPSC depression, also blocked the E139-induced EPSC depression. Furthermore, yohimbine, an alpha(2)-adrenoceptor antagonist, also blocked the E139-induced EPSC depression, while prazosin, a selective alpha(1)-adrenergic antagonist, and propranolol, a non-selective beta-adrenoceptor antagonist, did not block the E139 effect. Similar to the E139-induced EPSC depression, the NE-induced EPSC depression was also blocked by the GABA(B) receptor antagonist, CGP55845. By contrast, however, neither SCH23390 nor sulpiride, D1-like and D2-like DA receptor antagonists, respectively, blocked the E139-induced synaptic depression. These results suggest that NE and E139, but not DA, employ a similar mechanism to depress EPSCs in the NAc, and support the hypothesis that E139, like NE, may act on alpha(2)-adrenoceptors to cause the release of GABA, which then mediates synaptic depression via GABA(B) receptors.     

Neurosteroid pregnenolone sulfate enhances glutamatergic synaptic transmission by facilitating presynaptic calcium currents at the calyx of Held of immature rats

Pregnenolone sulfate (PREGS) is an endogenous neurosteroid widely released from neurons in the brain, and is thought to play a memory-enhancing role. At excitatory synapses PREGS facilitates transmitter release, but the underlying mechanism is not known. We addressed this issue at the calyx of Held in rat brainstem slices, where direct whole-cell recordings from giant nerve terminals are feasible. PREGS potentiated nerve-evoked excitatory postsynaptic currents (EPSCs) without affecting the amplitude of miniature EPSCs, suggesting that its site of action is presynaptic. In whole-cell recordings from calyceal nerve terminals, PREGS facilitated Ca2+ currents, by accelerating their activation kinetics and shifting the half-activation voltage toward negative potentials. PREGS had no effect on presynaptic K+ currents, resting conductance or action potential waveforms. In simultaneous pre- and postsynaptic recordings, PREGS did not change the relationship between presynaptic Ca2+ influx and EPSCs, suggesting that exocytotic machinery downstream of Ca2+ influx is not involved in its effect. PREGS facilitated Ba2+ currents recorded from nerve terminals and also from HEK 293 cells expressed with recombinant N- or P/Q-type Ca2+ channels, suggesting that PREGS-induced facilitation of voltage-gated Ca2+ channels (VGCCs) is neither Ca2+ dependent nor VGCC-type specific. The PREGS-induced VGCC facilitation was blocked by the PREGS scavenger (2-hydroxypropyl)-beta-cyclodextrin applied from outside, but not from inside, of nerve terminals. We conclude that PREGS facilitates VGCCs in presynaptic terminals by acting from outside, thereby enhancing transmitter release. We propose that PREGS may directly modulate VGCCs acting on their extracellular domain.     

Schizophrenia, hypocretin (orexin), and the thalamocortical activating system

Diminished connectivity between midline-intralaminar thalamic nuclei and prefrontal cortex has been suggested to contribute to cognitive deficits that are detectable even in early stages of schizophrenia. The midline-intralaminar relay cells comprise the final link in the ascending arousal pathway and are selectively excited by the wake-promoting peptides hypocretin 1 and 2 (orexin A and B). This excitation occurs both at the level of the relay cell bodies and their axon terminals within prefrontal cortex. In rat brain slices, the release of glutamate from midline-intralaminar thalamocortical terminals induces excitatory postsynaptic currents (EPSCs) in layer V pyramidal cells in prefrontal cortex. When hypocretin is infused into medial prefrontal cortex of behaving animals, it improves performance in a complex cognitive task requiring divided attention. Chronic restraint stress causes atrophy of the apical dendritic arbors in layer V prefrontal pyramidal cells and leads to a reduction in hypocretin-induced EPSCs, indicating impairment in excitatory thalamocortical transmission. Thus, taken together with evidence for an underlying loss of excitatory thalamocortical connectivity in schizophrenia, stress in this illness could further exacerbate a breakdown in cortical processing of incoming information from the ascending arousal system.     

The synaptic connectivity that underlies the noxious transmission and modulation within the superficial dorsal horn of the spinal cord

Noxious stimuli can usually cause the aversive sensations, pain and itch. The initial integration of such noxious information occurs in the superficial dorsal horn of the spinal cord (SDH), which is very important for understanding pain sensation and developing effective analgesic strategies. The circuits formed by pools of neurons and terminals within SDH are accepted as the platform for such complicated integrations and are highly plastic under conditions of inflammatory or neuropathic pain. Recent literature offers a complicated, yet versatile view of SDH intrinsic circuits with both inhibitory and excitatory components. However, our knowledge about the adaptative regulation of SDH local circuits is still far from sufficient due to the incomplete understanding of their organization as they are intermingled with primary afferent fibers (PAFs), poorly understood or identified SDH neurons, somehow contradictory data for descending control systems. A more positive view emphasizes abundant modern data on SDH neuron morphology and physiology riding on the back of significant technological advancements used in neuroscience. Reviewing the current literature on this topic thus produced an integrated understanding of SDH neurons and the SDH local circuits involved in noxious transmission and modulation.